In lignocellulosic biomass, crystalline cellulose fibrils are embedded in a less well-organized hemicellulose matrix which, in turn, is surrounded by an outer lignin seal. Contacting naturally occurring cellulosic materials with hydrolyzing enzymes generally results in cellulose hydrolysis yields that are less than 20% of theoretically predicted results. Hence, some “pretreatment” of the biomass is invariably carried out prior to attempting the enzymatic hydrolysis of the polysaccharides (cellulose and hemicellulose) in the biomass. Pretreatment refers to a process that converts lignocellulosic biomass from its native form, in which it is recalcitrant to cellulase enzyme systems, into a form for which cellulose hydrolysis is effective. Compared to untreated biomass, effectively pretreated lignocellulosic materials are characterized by an increased surface area (porosity) accessible to cellulase enzymes, and solubilization or redistribution of lignin. Increased porosity results mainly from a combination of disruption of cellulose crystallinity, hemicellulose disruption/solubilization, and lignin redistribution and/or solubilization. The relative effectiveness in accomplishing some (or all) of these factors differs greatly among different existing pretreatment processes. These include dilute acid, steam explosion, hydrothermal processes, “organosolv” processes involving organic solvents in an aqueous medium, ammonia fiber explosion (AFEX), strong alkali processes using a base such as, ammonia, NaOH or lime, and highly-concentrated phosphoric acid treatment. Many of these methods do not disrupt cellulose crystallinity, an attribute vital to achieving rapid cellulose digestibility. Also, some of these methods are not amenable for “easy recovery” of the chemicals employed in the pretreatment.
Of the existing pretreatment technologies, a coordinated development of the leading ones was recently reported (C. E. Wyman et al, Bioresource Technology, (2005) 96, 1959). A consortium of researchers (Biomass Refining Consortium for Applied Fundamental and Innovation (CAFI)) have studied the pretreatment of a well-characterized single feed stock, namely corn stover, by several then promising pretreatment technologies using common analytical methods, and a consistent approach to data interpretation (C. E. Wyman et al., Bioresource Technology, (2005) 96, 2026). In particular, the following were investigated: (1) dilute acid hydrolysis (T. A. Lloyd and C. E Wyman, Bioresource Technology, (2005) 96, 1967), (2) Ammonia Fiber Explosion (AFEX) technique (F. Teymouri et al., Bioresource Technology, (2005) 96, 2014), (3) pH Controlled liquid hot water treatment (N. Mosier et al., Bioresource Technology, (2005) 96, 1986), (4) aqueous ammonia recycle process (ARP) (T. H. Kim and Y. Y. Lee, Bioresource Technology, (2005) 96, 2007), and (5) lime pretreatment (S. Kim and M. T. Holzapple, Bioresource Technology, 96, (2005) 1994) were investigated. All of the abovementioned methods have two steps: a pretreatment step that leads to a wash stream, and an enzymatic hydrolysis step of pretreated-biomass that produces a hydrolyzate stream (FIG. 2). The combined total amounts of five and six carbon sugars and their oligomers in both these effluent streams were used to estimate the overall sugar yields in each of these methods.
In the above methods, the pH at which the pretreatment step is carried out increases progressively from dilute acid hydrolysis to hot water pretreatment to alkaline reagent based methods (AFEX, ARP, and lime pretreatments). Dilute acid and hot water treatment methods solubilize mostly hemicellulose, whereas methods employing alkaline reagents remove most lignin during the pretreatment step. As a result, the wash stream from the pretreatment step in the former methods contains mostly hemicellulose-based sugars, whereas this stream has mostly lignin for the high-pH methods. The subsequent enzymatic hydrolysis of the residual biomass leads to mixed sugars (C5 and C6) in the alkali based pretreatment methods, while glucose is the major product in the hydrolyzate from the low and neutral pH methods. The enzymatic digestibility of the residual biomass is somewhat better for the high-pH methods due to the removal of lignin that can interfere with the accessibility of cellulase enzyme to cellulose. All these methods are carried out in aqueous media at temperatures well above the normal boiling point of water to facilitate the physico-chemical phenomena involved in the depolymerization/melting of hemicellulose and lignin and require a high pressure environment (6 to 20 atm). Also, none of these methods can effectively disrupt the crystallinity of cellulose in the biomass. Furthermore, some of these methods release most of the xylose in the form of xylooligosaccharides, which are not easily fermented by many microorganisms and require additional steps to break them into monomeric species. Finally, a comparative economic analysis of these leading pretreatment technologies (T. Eggeman and R. T. Elander, Bioresource Technology, (2005) 96, 2019) indicates that they are all capital intensive compared to corn dry mill and significant process improvements are necessary for commercialization.
Organosolv methods in which solvents such as ethanol and methanol are used (in aqueous media) and methods using bases such as NaOH are also suggested in the literature. These methods are also capable of dissolving lignin, but they are not able to disrupt the crystallinity of cellulose. Further, the costs are so high that these methods are not considered competitive for manufacture of high-volume, low-value commodity products. Based on the information available on the concentrated phosphoric acid method (Zhang, Y-H P et al, Biotechnol. Bioeng. 97:214-223), it appears to be an elaborate method involving several steps making the overall process expensive. However, due to the harsh conditions used, amorphous cellulose is obtained by this pretreatment process.
Dissolution and processing of pure cellulose using ionic liquids was reported earlier (Swatloski, R. P., Rogers, R. D., Holbrey, J. D., U.S. Pat. No. 6,824,599, 2002; Holbrey, J. D., Spear, S. K., Turner, M. B., Swatloski, R. P., Rogers, R. D., U.S. Pat. No. 6,808,557, 2003). In a recent work, we reported on an effective approach to mitigate the recalcitrance of cellulose to enzymatic hydrolysis by Ionic liquid pretreatment (Dadi, A., Schall, C. A., Varanasi, S., “ ”, Applied Biochemistry and Biotechnology, vol. 136-140, p 407, 2007; Varanasi, S., Schall, C., and Dadi, A., US patent filed: February 2007). While this approach makes use of an ionic liquid to open the structure of pure crystalline cellulose material such as Avicel to make it accessible to cellulase enzymes, it did not specifically address the pretreatment of lignocellulosic biomass, which is the subject of this invention. In this context, it is noted that very recently isolation of cellulose from biomass by using ionic liquids (Fort, D. A., Remsing, R. C., Swatloski, R. P., Moyna, P., Moyna, G., Rogers, R. D., Green Chemistry 9: 63-69, 2007) and complete dissolution of biomass in ionic liquids (Vesa, M. Aksela, R., European Patent WO2005017001, 2005) have been investigated. The former application focuses on “in tact recovery” of cellulose portion of the biomass for materials development, whereas the latter aims to identify ILs and conditions that will lead to a “total dissolution” of biomass. However, currently a high yield pretreatment approach that can be used to rapidly and efficiently saccharify the polysaccharide portions of lignocellulosic biomass is unavailable. This invention exploits the differing “affinities” of the three major components of biomass (i.e., lignin, hemicellulose and cellulose) towards ILs, coupled with the unique capability of some ILs in disrupting the crystallinity of the cellulose portion (by breaking the hydrogen-bonding structure), to devise a scheme to efficiently saccharify the polysaccharide portions of biomass. The invention requires neither the extraction of cellulose from biomass nor the dissolution of biomass in IL.